The effector molecules of the immune system include a repertoire of circulating immunoglobulins non-attributable to exogenous antigenic induction, variously referred to as xe2x80x9cautoantibodiesxe2x80x9d or xe2x80x9cnatural antibodiesxe2x80x9d. The two terms are not synonymous. Thus, for the xe2x80x9cself-attackingxe2x80x9d antibodies the term xe2x80x9cautoantibodiesxe2x80x9d is customarily applied, while for the xe2x80x9cself-protectivexe2x80x9d antibodies the term xe2x80x9cnatural antibodiesxe2x80x9d is used.
A vast majority of natural antibodies react with one or more xe2x80x9cselfxe2x80x9d antigens. Their importance in immune regulation has long been neglected, since tolerance to xe2x80x9cselfxe2x80x9d was thought to be primarily dependent on the deletion of autoreactive clones, rather than on peripheral suppressive mechanisms. Clonal deletion cannot account, however, for the prevalence of natural autoreactivity among healthy individuals. It is now well established that autoreactive repertoires are predominantly selected early in ontogeny and that autoreactive antibodies, B cells, and T cells, are present in healthy individuals, and in virtually all vertebrate species (Lactoix-Desmazes et al., 1998, J. Immunol. Methods, 216:117-137 and references therein).
Natural antibodies are mostly IgM, polyreactive, and are generally encoded by V genes in germline configuration (Casali et al., 1996, Curr. Top. Microbiol. Immunol., 210:167-79 and references therein). They are mainly produced by B-1 cells which account for most of the B cell repertoire in the fetus and neonate, and possibly play a major role in the development of the adult B cell repertoire.
It is still unclear whether precursors of B-1 cells are capable of undergoing an antigen-driven clonal selection process, thereby producing natural antibodies with a high affinity for the selecting antigen. In this respect, it has been clearly established that B-1 cells can express a hypermutation mechanism similar to that of conventional (B-2) cells and that the main structural correlate for antibody polyreactivity and antigen binding in monoreactive antibodies is provided by the somatically generated CDR3 heavy chain (Casali et al., supra).
Although endowed with self-reactivity, natural antibodies also bind exogenous antigens. Exposure to environmental antigens is not necessary for the emergence of natural antibody-producing cell precursor clones to exogenous antigens, as suggested by the significant population of B cells capable of producing antibodies to a variety of bacterial antigens in germ-free animals (Casali et al., supra). Because of their ability to bind a variety of exogenous antigens, including those on bacteria and viruses, natural antibodies play a major role in the primary line of defense against infections.
U.S. Pat. No. 5,872,012 discloses a circulating natural human antibody immunoreactive with an arginine-rich epitope present on human protamine. U.S. Pat. No. 5,606,026 discloses that the arginine-rich epitope is present in the Tat protein of HIV-1 and further discloses a second circulating human natural antibody immunoreactive with a different epitope on the Tat protein. It has been also shown that these Tat-reactive circulating human natural antibodies decrease after HIV infection reaching minimal levels as the patient progresses to AIDS (Rodman et al., 1999, Hum. Immunol., 60:631-639). In addition, a novel circulating human natural antibody immunoreactive with a cryptic epitope present on human lactoferrin is disclosed therein.
As the correlation of the titers of some of the circulating natural antibodies with disease progression has been established in HIV infection, there is a need in the art to develop new treatment strategies based on supplementing the patient""s immune system with effective amounts of exogenously produced natural antibodies. An ideal source of such natural antibodies would be monoclonal counterparts of the circulating human natural antibodies that can be produced in large quantities and used for various therapeutic and diagnostic purposes.
The present invention provides monoclonal forms of human natural antibodies.
In one aspect, the present invention provides a method for producing human hybridoma cells producing monoclonal human natural antibodies comprising the steps of
fusing immortalized or transformed human umbillicord blood cells with mouse: human
heteromyeloma cells,
isolating fused cells,
plating said fused cells under limited dilution conditions, and
recovering said hybridoma cells.
In another aspect, the present invention provides human hybridoma cells producing monoclonal human natural antibodies produced by the method of the present invention.
These and other aspects of the present invention will be apparent to those of ordinary skill in the art in light of the present description, claims and drawings.